Ultrasonic synthesis, characterization, DFT and molecular docking of a biocompatible Zn-based MOF as a potential antimicrobial, anti-inflammatory and antitumor agent

Zinc metal–organic frameworks have emerged as promising candidates, demonstrating excellent biological properties stemming from the unique characteristics of MOFs and zinc. In this study, we employed a facile method to synthesize a zinc metal–organic framework [Zn(IP)(H2O)] using ultrasound irradiation, with the linker being isophthalic acid (IPA) (1,3-benzene dicarboxylic acid). The parent Zn-MOF and two Ag/Zn-MOF samples prepared via loading and encapsulation methods were comprehensively characterized using various techniques, including FT-IR, XRD, SEM, TEM, N2 adsorption–desorption isotherm, UV–vis spectroscopy and TGA. The parent Zn-MOF and two Ag/Zn-MOF samples exhibited a broad spectrum of antibacterial effects. Remarkably, genomic DNA of P. aeruginosa was effectively degraded by Zn-MOF, further supporting its potent antibacterial results. The free radical inhibition assay demonstrated a 71.0% inhibition under the influence of Zn-MOF. In vitro cytotoxicity activity of Zn-MOF against HepG-2 and Caco-2 cell lines revealed differential cytotoxic effects, with higher cytotoxicity against Caco-2 as explored from the IC50 values. This cytotoxicity was supported by the high binding affinity of Zn-MOF to CT-DNA. Importantly, the non-toxic property of Zn-MOF was confirmed through its lack of cytotoxic effects against normal lung cell (Wi-38). The anti-inflammatory treatment of Zn-MOF achieved 75.0% efficiency relative to the standard Ibuprofen drug. DFT and docking provided insights into the geometric stability of Zn-MOF and its interaction with active amino acids within selected proteins associated with the investigated diseases. Finally, the synthesized Zn-MOF shows promise for applications in cancer treatment, chemoprevention, and particularly antibacterial purposes.

alternatives to conventional antibacterial agents 5 .Zinc, in particular, plays a crucial role in various biological processes within the human body, serving as an enzymatic cofactor, signaling molecule, and structural element 6 .Additionally, zinc ions contribute to the regulation of Cu/Zn superoxide dismutase (SOD) activity 7 , indirectly modulating intracellular reactive oxygen species (ROS) scavenging.Research indicates a correlation between zinc depletion, decreased Cu/Zn SOD activity, and inflammation 8 , suggesting that exogenous zinc supplementation could potentially alleviate inflammation by restoring SOD activity.Moreover, zinc ions exhibit selective toxicity against cancer cells by inducing DNA damage, ultimately triggering cellular apoptosis.Consequently, zinc is being actively investigated as a therapeutic agent for tumor treatment 9 .
New porous compounds known as metal-organic frameworks (MOFs) have attracted increasing attention in recent years for their potential applications as antibacterial and anticancer materials.Additionally, they serve as a reservoir of metal ions 10 .These compounds exhibit structural characteristics containing cavities in the form of pores and channels, typically possessing high surface area, adjustable shape, and uniform-high pore volumes 11 .MOFs can effectively store metal ions, such as silver (Ag + ), nickel (Ni 2+ ), or copper (Cu + or Cu 2+ ) thereby demonstrating superior antimicrobial efficiency.Therefore, MOFs play a crucial role in inhibiting bacterial growth by gradually releasing metal ions through the degradation of framework structures, ensuring sustained antibacterial effects with high durability 12 .
While exposure to large amounts of Zn 2+ in a short period can be toxic 13 , the relatively stable frame structure of zinc-based MOF enables slow degradation in aqueous environments.This gradual degradation of MOF materials facilitates the long-term release of metal ions, achieving low toxicity levels 14 .Therefore, MOF particles have been shown to exhibit low cytotoxicity, enhancing their application as antibacterial agents relative to the other common disinfectants 1 .It is known that zinc compounds are extensively used in cosmetics and skincare, making them ideal candidates for cancer chemoprevention or as associated agents in clinical treatment 15 .An ideal anticancer agent would exert minimize adverse effects on normal tissues while maximizing its capacity to kill tumor cells and/or inhibit tumor growth 16 .Therefore, MOFs bearing zinc ions hold promise as suitable therapeutic agent for both bacterial and cancer therapies.
Traditionally, MOFs are prepared via room temperature stirring or solvothermal routes at high temperatures with extended reaction times.However, alternative techniques, such as microwaves 17 , electrochemistry 18 and mechanochemistry 19 are also employed.Ultrasounds irradiation presents another viable method for MOF synthesis, exploiting its high energy to reduce reaction times and enhance energy efficiency compared to other techniques like electrical heating or microwaves 20 .
A limited number of studies have highlighted the potential of Zn-MOF as potent antibacterial materials.For instance, Diéguez's et al. 21demonstrated the powerful antibacterial activity of MOFs based on zinc skeleton.Building on this foundation, our study aimed to explore the potential of Zn-MOF as a drug candidate.On the other hand, other studies have shown that the synthesized starched AgNPs, even at low concentrations, effectively inhibited the growth of various organisms as reported by Jansirani et al. 22 .Furthermore, Darabpour et al. 23 supported the concept of enhancing antimicrobial activity through the incorporation of AgNPs into Zn-MOF.Therefore, we anticipated an enhancement in the antibacterial behavior of the synthesized Zn-MOF upon incorporation with AgNPs.AgNPs were introduced through loading (L-Ag/Zn-MOF) and encapsulated procedures (C-Ag/Zn-MOF).
Considering the promising biological effects of zinc-based organic frameworks, we aim to report on the biological screening of Zn-MOF composed of the isophthalic acid linker, renowned for its biological activity 24 .Ultrasound irradiation is utilized for faster and environmentally friendly synthesis, resulting in reduced reaction time and energy consumption.Furthermore, considering silver's potential as an antibacterial agent and the observed good surface area of the MOF structures, efforts have been made to evaluate the concurrent antibacterial activity post-silver loading and encapsulation.Bacterial investigation includes the assessment of bacterial DNA damage, while cytotoxicity study extends to several cell lines.Moreover, the cytotoxicity study of Zn-MOF involves evaluating double-strand DNA binding and cleavage, along with exploring anti-inflammatory and antioxidant efficacy.Finally, DFT and molecular docking are performed to assess the geometric stability of Zn-MOF and its interaction with active amino acids within selected proteins associated with the investigated diseases.

Experimental Materials and physical measurements
All chemicals and solvents, including zinc acetate dihydrate, isophthalic acid (IPA) and ethanol, were of pure analytical grade and obtained commercially from Sigma-Aldrich Company.The synthesized [Zn(IP)(H 2 O)] and two Ag/Zn-MOF samples prepared via loading and encapsulation methods were characterized through various techniques using TGA, SEM, TEM, XRD, FTIR, UV-vis spectroscopy and BET surface area analysis.
FT-IR spectra were recorded using a Shimadzu FT-IR spectrometer within the range 400-4000 cm −1 .Electronic absorption spectra in DMF were measured utilizing an automated UV/Vis-NIR 3101 PC Shimadzu spectrophotometer.
The thermal measurements were performed using a TGA-50H-Shimadzu thermal analyzer under a nitrogen atmosphere, heating from room temperature to 800 ℃ at a rate of 10 ℃ min −1 .X-ray diffraction analysis involved irradiating the sample with monochromatic X-rays across a range of incident angles.The angles and intensities of the X-ray diffraction were measured using a Bruker LynxEye detector.
The micro-structural morphology was investigated via Scanning electron microscopy (SEM) using a Quanta 250 FEG instrument.The investigation of particle size was conducted using a transmission electron microscope (TEM), specifically a model Jeol JEM 1400 manufactured in Japan, operating at 80 kV.Additionally, specific surface area was obtained utilizing BET analysis with N 2 adsorption/desorption at 77 K, performed using a gas

Well diffusion method
The antibacterial and antifungal effectiveness of synthesized Zn-MOF and its hybrid forms, Ag loaded and encapsulated, were evaluated against Bacillus Subtilis, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Candida albicans, Aspergillus Niger microorganisms.The in vitro antimicrobial activities of the synthesized compounds were assessed using a modified agar well diffusion method 25 .After incubation at 35-37 ℃ for 24-48 h with scanned strains, the diameters of the inhibition zones were measured in millimeters.The growth of the tested strains reached approximately 10 7 cells/mL using Mueller Hinton media and quantified using a plate counter.A volume of 100 μL of microbial suspension was spread onto agar plates.Subsequently, the volume 10 μL of the tested samples was placed on agar media using blank paper disks.

P. aeruginosa DNA fragmentation
P. aeruginosa bacteria were cultured at 10 5 CFUs/mL in Luria Bertani (LB) broth-rich medium in the presence and absence of the tested compounds at different concentrations ranging from 5 to 25 μg/mL.The cultures were incubated for 4-5 h at 37 ℃ with shaking at 120 rpm in an incubator shaker 26 .Subsequently, the growing bacteria were centrifuged at 8000 rpm for 10 min and washed twice with Phosphate Buffer Solution (PBS) to remove residual media.
Genomic DNA was isolated using the Gene JET Genomic DNA Purification Kit (Thermo Scientific) to assess the genomic DNA integrity.The extracted DNA was evaluated for its integrity using agarose gel electrophoresis.DNA samples were electrophoresed on a 1.0% agarose gel in Tris Boric EDTA (TBE) buffer for 45 min at 90 V.The gel was then stained with ethidium bromide (EtBr), and an image was captured using a mobile camera.

Antioxidant and antitumor study
Free radical scavenging activity was measured by 1,1-diphenyl-2-picryl hydrazyl (DPPH) assay.Initially, a 0.1 mM solution of DPPH in ethanol was prepared.Subsequently, 1 mL of this solution was added to 3 mL of different concentrations (3.9, 7.8, 15.62, 31.25,62.5, 125, 250, 500, 1000 μg/mL) of [Zn(IP)(H 2 O)] in DMSO.The mixture was vigorously shaken and allowed to stand at room temperature for 30 min, then absorbance was measured at 517 nm using a spectrophotometer.Ascorbic acid was used as the reference standard compound, and the experiment was done in triplicate 27 .The IC 50 value of the tested sample was calculated.The percentage DPPH scavenging effect was calculated using the following equation: where A 0 represents the absorbance of the control reaction and A S represents the absorbance in the presence of a test or standard sample.
The cytotoxic effect of [Zn(IP)(H 2 O)] was determined by using a 3-(4,5-dimethylthiazolyl)2,5-diphenyltetrazolium bromide (MTT) assay 28 .Initially, each cell line, including HepG2, Caco2 and Wi-38 was inoculated in a flat-bottomed 96-well plate at a density of 10 4 cells/100 µL complete RPMI medium/well and allowed to develop a complete monolayer sheet overnight.After 24 h, the cell monolayer sheet was washed twice with wash media and treated with a two-fold dilution series of the tested samples ranging from 0.0 to 1000 µg/mL.The dilutions were achieved by dissolving the proper weight of the tested sample in 1 mL of PRMI medium containing 2% fetal bovine serum (FBS).Subsequently, 100 µL of each diluted sample was loaded in different cultured wells, and the experiment performed in triplicate.Three wells containing inoculated cells received culture medium and served as untreated groups.The plate was then incubated for 24 h in a humidified incubator at 37 ℃ with 5% CO 2 .After that, cells were examined under an inverted microscope to observe any signs of toxicity induced by [Zn(IP) (H 2 O)] on each cancer cell line.MTT solution (BIO BASIC CANADA INC) (20 µL of 5 mg/mL PBS) was added to each well, and the plate was shaken at 150 rpm for 5 min to ensure thorough mixing.Subsequently, the plate was returned to the incubator to allow MTT to metabolize and appear as purple formazan crystals.After incubation, the medium was dumped off and the formazan crystals were dissolved in 200 µL of DMSO by shaking the plate on the shaking table for 5 min at 150 rpm.The optical densities (OD s ) of the dissolved formazan crystals

DNA cleavage
The impact of the prepared [Zn(IP)(H 2 O)] on the DNA cleavage of PBR322 (100 ng) was evaluated using concentrations ranging from 5 to 20 μg/mL in a final reaction mixture of 10 μL in the presence of control.Each reaction was then incubated in darkness at 37 ℃ for 1 h.To halt the reaction, 6 × loading dye was added, and the samples were monitored by running them on 1.0% agarose gel in TBE buffer at 70 V for 45 s, then the gel was stained by EB and photographed using mobile camera.

Denaturation inhibition measurements of bovine serum albumin
In the protein denaturation inhibitor screening, a modified method of Gambhire et al. was employed 29 .In a reaction volume of

Computational study
The calculations were performed using Gaussian 09 revision A.02 30

FTIR spectra
The FTIR spectrum of the synthesized Zn-MOF sample revealed characteristic features (Fig. 1).The O -H stretching modes of water molecules typically appear above 3000 cm −1 .The FTIR spectrum of Zn-MOF exhibits two bands at 3415 and 3192 cm −1 , which are assigned to the antisymmetric and symmetric OH stretching modes, respectively, of coordinated water molecules 31 .The spectrum of IPA exhibits a strong and broad band centered around 3000 cm −1 which corresponding to (O-H) vibrations of hydrogen bonds.This is typical for carboxylic acids, which usually exist as hydrogen-bonded dimers 32 .The absence of an absorption band around 3000 cm −1 in the spectrum of the Zn-MOF confirmed the full deprotonation of the IPA linker during the formation of the Zn-MOF.
The IR bands corresponding to the ν(C = O) and ν(C -O) modes of the IPA are reported at ca. 1690 and 1280 cm −1 , respectively 33,34 .However, in the Zn-MOF, these absorption bands are replaced by the ν(COO − ) of carboxylates with C -O bond 31 .Moreover, the C -O stretching modes of the carboxylate groups are identified by the presence of antisymmetric and symmetric vibrations, ν as (COO − ) at ca. 1600, 1532 and ν s (COO − ) at ca. 1462, 1408 cm −1 , respectively.The difference in wavenumbers between these two frequencies {Δ = ν as (COO − ) − ν s (COO − )} provides information on the binding mode of the carboxylate groups to zinc metal centers.Typically, values of Δν exp above 180 cm −1 are indicative of monodentate coordination, whereas values of Δν exp below 120 cm −1 usually suggest chelating or bridging carboxylate groups 31,35 .For Zn-MOF, the experimental values of Δν exp are 192 and 70 cm −1 , corresponding to monodentate and bridging (or chelating) binding mode, respectively.However, it is essential to note that these correlations are empirical and cannot be used for a definitive structure.Moreover, the O -H modes of IPA are replaced by M -O modes, observed at low frequencies 31 , with bands appearing at 472 and 469 cm −1 assigned to Zn -O 36,37 .These analyses suggested that IPA was successfully converted to Zn-MOF, which presents one fourfold-coordinated zinc ion with two bridging oxygen and one monodentate from carboxylate linker, along with one additional coordinated water molecule, i.

e. [Zn(IP)(H 2 O)].
The FT-IR spectra of the Ag/Zn-MOF did not show significant differences compared to the spectrum of Zn-MOF alone, with changes primarily observed in peak intensities rather than positions.In general, the similarity observed suggests that the coordination states of the components remain unchanged following silver incorporation 38 .

Uv-vis spectra
The transitions corresponding to the bands of the IPA ligand and its Zn-MOF were represented in Fig. 2. IPA showed transitions at 279.0 and 288.0 nm, which can be assigned to π-π* transitions, specifically involving the aromatic part, and n-π* transitions of the non-bonding electrons of (C = O), respectively.Upon bonding formation, the transition spectrum of the MOF observed at 277.0 and 341.0 nm.Since Zn(II) ions have a d 10 electron configuration and are difficult to oxidize or reduce, the transitions observed are probably attributed to intraligand π -π* transitions, perturbed by metal coordination 39 .In Zn(II) carboxylate-based compounds, the HOMO and LUMO are likely the π-bonding orbital from the aromatic rings and the Zn -O (carboxylate) π*-antibonding orbital, respectively, often localized on the metal centers [37][38][39][40] .The d 10 configuration and its diamagnetic properties Zn(II) ions suggest a possible tetrahedral geometry.The aqueous silver ions were reduced to silver nanoparticles (AgNPs), resulting in a brownish-yellow color, indicative of AgNPs formation, as illustrated in the experimental section.The formation and stability of these nanoparticles were examined using UV-Vis spectrophotometry, with the maximum absorbance observed at 438 nm, consistent with previous findings 22,41 .
The successful attachment and incorporation of the AgNPs onto the Zn-MOF are highlighted in the UV-Vis experiments, as seen in Fig. 2. The UV-vis spectra of Ag/Zn-MOF samples retained the characteristic features of Zn-MOF, exhibiting a narrow and sharp excitation wavelength in the range of 283-311 nm, while the presence of AgNPs was evident at 438 and 440 nm for L-Ag/Zn-MOF and C-Ag/Zn-MOF, respectively.

Thermal analysis
Figure 3 shows the thermogram of the synthesized ultrasonic-assisted [Zn(IP)(H 2 O)] sample under the optimized inert condition.Thermal behavior of the compound is characterized by two distinct regions.In the first region, observed at 170.0 ℃, there is a loss of the coordinated water molecule directly bonded to the zinc ion, resulting in a reduction in initial mass with a loss percentage of 7.66% (calc.= 7.27%).The second step involves the loss of the organic moiety (isophthalate) at 457.3 ℃, with a weight drop of 59.68% (calc.= 59.84%).This step leaves behind ZnO residue, with a mass of 32.66% (calc.= 32.88%).For the two Ag/Zn-MOF samples, a similar decomposition behavior to [Zn(IP)(H 2 O)] was observed, as seen in Fig. 3.The decomposition occurred in two main weight loss steps.For L-Ag/Zn-MOF and C-Ag/Zn-MOF, the second weight loss step occurred at relatively higher temperatures (474 and 467 ℃, respectively) compared to Zn-MOF, indicating the improved thermo-stability of the MOF after AgNPs incorporation.The presence of silver nanoparticles in the structures leads to higher residual percentage values compared to pristine MOF, as the final residues comprise not only zinc oxide but also silver metal.This results in 36.1 and 35.0%residual precipitate for L-Ag/Zn-MOF and C-Ag/Zn-MOF, respectively.The estimated silver precipitate accounts for 3.4 and 2.3% relative to L-Ag/Zn-MOF and C-Ag/Zn-MOF, respectively.
Coats-Redfern (CR) and Horowitz-Metzger methods were used to evaluate the kinetic and thermodynamic parameters (ΔE, ΔH, ΔS, and ΔG) 40 , as seen in Fig. 3 and summarized in Table 1.The high values of the activation energy (ΔE*) can be related to the stability of the investigated Zn-MOF.The activation entropies ΔS* have small positive values which may indicate a small change in the ordered state of the activated compounds and the reactants 43 .Although the change is small, the positive value suggests an increase in disorder during the reaction.The positive values of ΔG* indicate the non-spontaneity of the processes.This implies that the processes are not favored in terms of spontaneity and require an input of energy to proceed.010), (012), ( 111), ( 200), ( 201), ( 104), ( 211), ( 006), (008), (217) reflections, respectively.Table 2

displays a comparison between the theoretical and experimental peaks positions of the prepared [Zn(IP)(H 2 O)].
The XRD characterization of the two Ag/Zn-MOF samples revealed that their crystal structures remained intact, with the main peaks observed at the same positions (Fig. 4).However, slight differences in peaks positions and intensities were observed, which could be due to changes in their morphology during synthesis.The Table 1.The activation parameters* for the decomposition of Zn-MOF using Coats-Redfern and Horowitz-Metzger methods respectively.

Compound
Step  The morphology, including size and shape, of the Zn-MOF material was analyzed using scanning electron microscopy (SEM) as shown in Fig. 5a.The SEM image showed aggregated polygonal plate-like morphology of Zn-MOF particles with varying dimensions.On other hand, the size and shape of the silver nanoparticles were analyzed using transmission electron microscopy (TEM) as depicted in Fig. 5b.Understanding the size distribution of particles is particularly crucial, especially for small-sized particles.The particle size distribution was assessed by measuring the diameter of Ag particles utilizing TEM images processed with ImageJ software, as shown in Fig. 5c.Subsequently, the obtained values were depicted in a histogram and fitted with a Gaussian function, revealing a mean particle size of less than 10 nm, as evidenced by the particle size distribution curve.These sizes are significantly smaller than typical bacteria, suggesting their potential efficacy as antimicrobial agents.
In the case of loaded form with AgNPs, the SEM image showed clear accumulation of AgNPs on the surface of the Zn-MOF and this observation was further supported by the measured surface area obtained from N 2 adsorption-desorption experiment, as seen in Fig. 6.The calculated BET-specific surface area of the modified Zn-MOF was higher than that of the pristine Zn-MOF indicating an increase in surface area due to the presence of AgNPs (cf.Table 3).However, the average pore size of the modified Zn-MOF was found to be lower than that of the pristine Zn-MOF.The increase in surface areas of Ag/Zn-MOF, particularly in the case of the loaded samples, is significant as it provides more available surface area for contacting microbial species.The increased surface area offers more active sites for adsorption and contact with the investigated microbial species, as compared to the parent Zn-MOF 45 .

DFT study
The energy value of the prepared [Zn(IP)(H 2 O)] was evaluated using the density functional theory (DFT), with an estimation of several parameters such as dipole moment, hardness (η), softness (S), energy gap, chemical potential (μ) and electronegativity (χ), as mentioned in Table 4.The structure numbering systems and the frontier molecular orbitals (FMO) of the (IPA linker and [Zn(IP)(H 2 O)]) were explored in Figs. 7, 8.The DFT analysis revealed that [Zn(IP)(H 2 O)] exhibited more negative values than the starting IPA ligand, indicating a favorable chelation process.The previous parameters were calculated using the following equations: [Zn(IP)(H 2 O)] was found to be a softer molecule than its parent IPA linker, depending on the HOMO-LUMO energy gap.A smaller gap indicates a softer molecule, with the HOMO and LUMO representing the FMO as shown in Fig. 8.The capacitance to accept electrons is called softness and vice versa with hardness 46 .
The energy needed to get rid of the electron is defined by the ionization potential, which equals the HOMO's energy whereas the LUMO's energy corresponds to the electron affinity, which produces energy when the system adds one additional electron 47 .A large energy difference was observed between the HOMO energy values of the IPA linker and the zinc-free ion; the linker exhibited a higher LUMO energy (− 1.45 eV) because it acted as the donating source of electrons and vice versa in the case of zinc-free ion (− 12.44 eV).Also, the water molecule, functioning as coordinated ligand, showed analogous behavior to IPA.The electron cloud of the IPA ligand's HOMO was mainly localized on the carboxylate group.However, in the case of [Zn(IP)(H 2 O)], the zinc ion became included with the carboxylate group.Similar electron density distribution was observed in the case of the LUMO, where both the carboxylate and zinc ion species played crucial roles in the coordination system of [Zn(IP)(H 2 O)].The negative value for chemical potential and high value for ionization potential contribute to the stability of the compound 48 .The polarity of the IPA linker is increased after binding with zinc ion, as evidenced from the magnitude of their dipole moments 49 .
Changes in bond lengths and angles were observed upon coordination (see Table 5 and Fig. 7), with some bonds elongated and others shortened to achieve the desired tetrahedral structure of the formed  The atomic charges were calculated using Mulliken and Natural Population analysis (MPA and NPA), revealed changes in charge distribution, with some atoms (O8, O9 and O11) exhibiting predominantly negative charges, particularly those involved in coordination with the zinc ion.The molecular electrostatic potential (MEP) analysis indicated specific structural reactive sites, with atoms exhibiting positive electrostatic potential can connect to the electrostatic negative potential sphere, serving as active sites for zinc ion coordination.
Overall, the investigated [Zn(IP)(H 2 O)] exhibited positive electrostatic potential (ESP) maps on their skeletons and negative ESP on their coordination centers, enhancing their antimicrobial activity 54 .The charge of (O11) as a representative example was changed from − 0.58 to − 0.46 due to the charge transfer from donating active atom to zinc ion with increasing the electron densities along the zinc ions from + 2 to + 0.519150, + 0.402530 and + 0.945793 as observed in Zn24, Zn23 and Zn20 repeated zinc ions, respectively.The oxygen electron density of the water molecule was decreased upon complexation from − 0.976 to − 0.674 corresponding to the uncoordinated and the coordinated water states respectively, this is due to ligand to zinc charge transfer (L → M), sp 3 hybridization binding type of the assumed tetrahedral structure.The Atomic charge changes for IPA and its Zn-MOF are tabulated in Table 6.

Molecular dynamics (MD) simulation
The theoretical study was extended to investigate the adsorption of silver atoms on zinc centers of [Zn(IP) (H 2 O)].Using the material studio program and adsorbate calculation, various planes (hkl) identified from the XRD chart were evaluated for modeling the adsorption process and determining the adsorption energy.Figure 9 and Table 7 represent different planes of Zn-MOF with calculated adsorption energy values relative to specific planes.The negative binding energies represent a stronger interaction between silver atoms and the zinc centers.Density of States (DOS) plots were analyzed to assess electronic band parentage, providing information about the type of charge transport that may exist 55 .Figure 9 shows the density of states, represented by the number of electrons.We found the order of electron density to be ( 104   be correlated to the number of available zinc atoms serving as the adsorbate surface for the silver atom in each selected plane.Table 7 presents the output and descriptors calculated by Monte Carlo simulation, including parameters such as substrate-adsorbate total energy, deformation energy and rigid adsorption energy.The substrate-adsorbate total energy reflects the sum of deformation and rigid adsorption energies, while the deformation energy reports the energy released when the adsorbate components are relaxed on the substrate surface Rigid adsorption energy refers to the energy released (or required) when the unrelaxed adsorbate components are adsorbed on the substrate.

Antibacterial and fungal activity with docking study
Agar well diffusion method Evaluation of the antibacterial effects against different bacterial strains was conducted using the agar well diffusion method, with inhibition zones serving as indicators of antimicrobial activity (Figs. 10, 11 and Table 8).
The results demonstrated a broad-spectrum inhibition effect against both gram-positive and gram-negative bacterial strains, including Bacillus Subtilis, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa.
Remarkably, the synthesized samples exhibited comparable or even superior activity to the standard antibiotic, Gentamicin, against gram-positive bacteria.The enhanced efficacy of the C-Ag/Zn-MOF sample, against B. Subtilis and S. aureus, may be attributed to the combined effects of AgNPs and [Zn(IP)(H 2 O)].The increased surface area of Ag/Zn-MOF, as determined from BET measurements, provides more active sites for interaction with bacterial species.Furthermore, the controlled release of their components from MOF, either as reservoirs or Table 5.Some of the optimized bond lengths, Å and bond angles, degrees, for IPA ligand and its Zn-MOF using B3LYP/6-311G and B3LYP/LANL2DZ, respectively.carriers, emphasizes their advantage in antimicrobial applications 56 .Silver nanoparticles, can permeate microbial cell membranes, disrupting DNA and enzyme function, leading to microbial destruction 57,58 .Interestingly, SEM images of Ag/Zn-MOF illustrate the adsorption of AgNPs (Fig. 6), further supporting the enhanced antibacterial activity observed.Notably, [Zn(IP)(H 2 O)] alone exhibited potent suppression of gram-negative growth, achieving inhibition percentages of 107.14 and 123.8% against E. coli & P. aeruginosa, respectively relative to the standard antibiotic used.This may be attributed to their highly negatively charged surface, possibly related to the lipopolysaccharides or teichoic acids with phosphate-rich constituents.Such surface characteristics may favor the release of positively charged zinc ion, which could act more effectively against gram-negative bacteria during the examination.However, the surface of AgNPs introduced through loading and encapsulating techniques provided zero-charged Ag particles.This characteristic may result in lower attraction with the negatively charged bacterial surface.Additionally, it may contribute to blocking the release of zinc ions as seen in the morphology structures of Ag/Zn-MOF observed in both cases.
Moreover, it was reported that concentrations above 10 −4 M of Zn 2+ ions could disturb cellular homeostasis and exert cytotoxic effects on prokaryotes 59 .In general, the varied effectiveness against different bacterial strains is likely influenced by their distinct cellular structures 60 .Unlike S. aureus, E. coli possesses an additional outer layer of liposaccharides, this structural difference can lead to differences in their functions and responses to antibacterial agents 61 .
Comparative analysis with other Zn-based MOFs revealed the superior antibacterial activity of our synthesized samples.Table 9 illustrates the more favorable effects observed in our samples.For instance, Zn-MOFs based on terephthalate, studied by Akhbari et al., reported a maximum inhibition zone value of 16 mm against S. aureus using 5000 μg/mL 1 .Additionally, both Zn-based materials studied by Diéguez et al. 21composed of 5-((4-carboxyphenyl)ethynyl), isophthalic acid, and formate anion, achieved significant antibacterial performance against both S. aureus and E. Coli strains, likely due to more progressive Zn release.Furthermore, Darabpour et al. 23 discussed the antibacterial results of a Zn-MOF based on 2-Aminoterephthalic acid and 1,4-bis(4pyridyl)-2,3-diaza-1,3-butadiene, combined with fabricated AgNPs/Zn-MOF nanocomposites achieved a 12 mm against the S. aureus strain at 8 mg/mL of the composite.In conclusion, our study highlights the potential of the synthesized [Zn(IP)(H 2 O)] as an effective antibacterial agent, surpassing standard market drug and similar Zn-MOFs.
Fungal species such as Candida albicans and Aspergillus Niger were tested against the three samples in comparison to the effect of the standard drug amphotericin B. Unfortunately, our compounds exhibited no activity against Aspergillus Niger.Nonetheless, the AgNPs loaded sample showed promising results against C. albicans outperforming the standard drug, while the parent compound [Zn(IP)(H 2 O)] exhibited similar inhibition activity to the reference drug.

Molecular docking
A molecular docking investigation was performed on the prepared [Zn(IP)(H 2 O)] against glucosamine-6-phosphate synthase enzyme (Code:1XFF).Various types of interaction with each amino acid were tabulated in Table 10 and were represented in Fig. 12, along with scoring energy and hydrogen bond values.The best-fit sit of enzyme interaction with [Zn(IP)(H 2 O)] revealed a backbone donor interaction between hetero atom source (O) from Gly-99 amino acid and the Zn-MOF.A backbone acceptor interaction was observed between the water Table 6.Atomic charges in terms of (MPA and NPA) for IPA and its Zn-MOF using B3LYP/6-311G and B3LYP/LANL2DZ, respectively.The MOF docking using the co-crystalized glutamate ligand binding site produces a more effective scoring energy with a more negative value of − 5.98 kcal/mol, indicating favorable interaction.This interaction involved different amino acids such as Thr-76, Gly-99, His-97, Trp-74 and Cys-1.The carbonyl oxygen of the isophthalate linker interacted with Thr-76 and Cys-1 amino acids, while the water molecule interacted with Gly-99, His-97, and Trp-74.Water molecule is the main source of hydrogen bond formation with bond lengths mentioned in Table 10.Gentamicin, when docked, exhibited a scoring energy value lower than [Zn(IP)(H 2 O)], consistent with its antibacterial results.Gentamicin interacted with Gly-99, His-86, Thr-76, Thr-124 and Asp-123 amino acids, with OH and -O-functional groups shared from the gentamicin molecule with different amino acids.The docking procedure was validated by re-docking the co-crystalized compound glutamic acid in its binding pocket,     11B).This resulted in smearing, corresponding to degraded genomic DNA compared to the intact one observed in the control (in the absence of [Zn(IP)(H 2 O)]).These findings suggest that the tested compound is strong enough to induce fragmentation of P. aeruginosa DNA, highlighting the potential of Zn-MOF as an antimicrobial agent.This result confirms the findings of the antimicrobial activity screening.

Structure-activity relationships of antioxidant and antitumor activity in vitro
Antioxidants perform by inhibiting the effects regulated via free radicals and oxidizing compounds.The evaluation of the free radical scavenging activity is a typically performed using DPPH assay, where DPPH molecule is neutralized upon accepting hydrogen or an electron 62 .The scavenging activity percentages of [Zn(IP)(H 2 O)] were calculated at different concentrations and compared to ascorbic acid, a positive standard drug.The IC 50 value, representing the concentration at which 50% of the DPPH radicals are scavenged, was obtained by plotting the scavenging activity percentages against the different sample concentrations.The loss of the DPPH transition signal intensity in the presence of our suggested antioxidant is directly proportional to the number of protons or electrons accepted.[Zn(IP)(H 2 O)] exhibited an IC 50 value of 66.07 µg/mL, compared to 55.5 µg/mL for the standard drug Vitamin C. Scheme 1 suggests that the coordinated water molecule acts as a proton source due to its induced ionization under the effect of coordination effect, facilitating the ionization of water molecule 63 .Metal chelation is speculated to substantially increase the dissociation constants of the acid substituent in the bound ligand, compared to the free ligand 64 .This interpretation aligns with the high complex formation ability of zinc-chelated compounds, consistent with the Irving-Williams's order for divalent metals of the 3d series 65 .In vivo studies have indicated that the enhanced antioxidant activity of Zn compounds may result from the acquisition of additional superoxide dismutating centers, increasing in the molecule's ability to stabilize unpaired electrons and scavenge free radicals according to Souza et al. 66 .The biological screening was extended to study the potential of [Zn(IP) (H 2 O)] as an antitumor agent.To evaluate the potential of [Zn(IP)(H 2 O)] against colony and liver cancer cells, a standard cell proliferation assay was performed.The inhibition effect versus concentrations is shown in Fig. 13, and images of the cell lines in case of control and treated states with various concentration of [Zn(IP)(H 2 O)] were captured (Figs.14, 15).The IC 50 values against human tumor Caco-2 colon and HepG-2 liver cell lines inhibitory concentration which provides 50% killing (Fig. 13) were observed, exhibiting earlier achievement of IC 50 against colon cancer cells compared to liver cancer cells.Specifically, [Zn(IP)(H 2 O)] achieved 50% killing of human tumor Caco-2 colon cell line at an inhibitory concentration of 79.83 µg/mL, with over 80% growth inhibition at 125 µg/mL.In contrast, the IC 50 value for HepG-2 liver cancer cells was 122.35 µg/mL, with 80.0% inhibition efficiency reached at ≈ 250 µg/mL.Therefore, it seems that the drug resistance in liver cancer cells is

Molecular docking
The antitumor docking study results are reported in Fig. 16 and Table 11 11).The docking results revealed the interaction between the (N1) quinazoline ring of AQ4 inhibitor and the terminal amino group of Lys-721 amino acid.All bond lengths and amino acids involved in the interactions are tabulated in Table 11.

DNA binding and cleavage studies
Covalent binding in DNA is irreversible and invariably leads to complete inhibition of DNA processes and subsequent cell death.Cis-platin (cis-diamminedichloroplatinum), a well-known covalent binder used in cancer therapy, forms intra/inter-strand cross-link via its chloro groups with nitrogen on the DNA bases 67 .Therefore, DNA binding was investigated to evaluate the suitability of [Zn(IP)(H 2 O)] as an effective anticancer drug.To evaluate the DNA binding affinity of [Zn(IP)(H 2 O)], different concentrations (5-25 µg/mL) of the Zn-MOF were used to monitor electrophoretic mobility and changes in bands intensity of CT-DNA in an agarose gel.The results showed changes in band intensity and migration of CT-DNA induced by binding with the tested compound relative to the control (free CT-DNA) (Fig. 17A).The decrease in electrophoretic mobility of DNA suggests a decrease in total charge and an increase in molecular mass due to binding with [Zn(IP)(H 2 O)] and/or change in the CT-DNA structural conformation 68 .The observed decrease in band intensity (Fig. 17A) with increasing Zn-MOF concentration may be attributed to several factors.Firstly, covalent binding of Zn-MOF with CT-DNA could damage parts of the DNA, affects its length visualization intensity by ethidium bromide stain 69 .Secondly, the steric effect induced by DNA binding to the metal in Zn-MOF may hinder ethidium bromide intercalations.Thirdly, the planar aromatic moiety of the [Zn(IP)(H 2 O)] could intercalate into adjacent DNA pairs, inhibiting ethidium bromide binding to CT-DNA.
The DNA cleavage pattern was further examined using circular DNA plasmid of PBR322.The results indicated that [Zn(IP)(H 2 O)] at 20 μg/mL could convert a relative amount of DNA to the supercoiled form (SC form) while depleting the open circular form (OC form) compared to control plasmid DNA (Fig. 17B).The OC form is produced by scissoring of the SC form 70 .The increase in supercoiling percentage suggests enhanced plasmid quality, which could be beneficial for gene therapy 71 .Therefore, [Zn(IP)(H 2 O)] promise for gene therapy by generating more supercoiled DNA vectors.

Anti-inflammatory activity
Protein denaturation activity Non-steroidal anti-inflammatory drugs (NSAIDs) treat inflammation by inhibiting of protein denaturation, inhibiting, hydrolysing enzymes or promoting membrane stabilization.In general, inflammation is caused by either lysosomal enzymes released by leukocytes or protein denaturation as in the case of arthritis 72 .To further explore the pharmacological and biological potential of [Zn(IP)(H 2 O)], its ability to inhibit protein denaturation inhibition was screened.The compound exhibited 50% ± 0.02 inhibition at 10 µg/mL (the obtained result is the mean of three individual measures) indicating its potential as a protein inhibitor and promising anti-inflammatory agent (Fig. 18).This efficacy is comparable to 75.0% efficiency relative to the standard drug ibuprofen.The positive charge of the released zinc ion interacts with BSA which carries a negative charge at physiological pH 73 .Thus, the protein has an overall negative charge (− 18e) at pH 7 74 .Also, the water molecule in [Zn(IP)(H 2 O)] can form hydrogen bonds with BSA.Although further studies are needed to fully understand and confirm the anti-inflammatory activity of [Zn(IP)(H 2 O)], these initial findings suggest its potential for treating inflammation.Finally, the synergistic effects of its antibacterial and anti-inflammatory properties suggest its potential application as a skin wound healing agent.

Figure 4
illustrates the XRD pattern of the prepared [Zn(IP)(H 2 O)].The sharp peaks observed in the pattern indicate its inherent crystallinity.The compatibility between the experimentally observed XRD pattern of the prepared Zn-MOF and the theoretical XRD pattern of the previously reported single crystal of
These observations are due to the formation of the (Zn -O = C) and (Zn -O -C = O) bonds, which make the carbonyl C = O bond weaker and accepted some of the single bond characters[50][51][52][53] .The tetrahedral structure was completed with the coordination of water molecule.
and the carboxylate group (O) of Trp-74 amino acid; with a hydrogen bond length of 2.08 Å.The scoring energy achieved was − 3.65 kcal/mol supporting the excellent experimental antibacterial results.

Fig. 11 .
Fig. 11.(A) Biological activity of Zn-MOF and its hybrid AgNPs towards different types of bacterial and fungal strains; (B) The DNA profile damage of P. aeruginosa in the absence (control) and presence of the tested drug (10 µg/mL) relative to control using 1% DNA gel electrophoresis.
higher than the colon cancer cells, or it can expect that the Caco-2 receptors exhibit greater affinity to [Zn(IP) (H 2 O)] compared to the HepG-2 receptors.It is desirable for a substance to show good cytotoxic activity against various human cancer cells with minimal or no effect on normal cells.So, the study was expanded to investigate the cytotoxic effect of [Zn(IP)(H 2 O)] on a human normal cell line using the human normal lung (Wi-38) cell line.[Zn(IP)(H 2 O)] exhibited minimal adverse effects on normal human lung (Wi-38) tissues.The observed IC 50 value of 225.79 µg/mL on the Wi-38 cell line suggests minimal activity of [Zn(IP)(H 2 O)] on normal lung cells.Considering that IC 50 values represent

Fig. 13 .
Fig. 13.In vitro cytotoxicity of Zn-MOF against the three investigated cell lines with the IC 50 values.
. The most important interaction is the formation of hydrogen bonds, especially those with bond lengths less than 3.0 Å.The most predominant types of interactions observed are the backbone and side chain acceptors.For instance, [Zn(IP)(H 2 O)] formed hydrogen bonds with Phe-771 (bond length 2.08 Å), while Glu-780 and pro-770 formed hydrogen bonds with the water molecule of [Zn(IP)(H 2 O)].The scoring energy value reveals the high binding affinity of [Zn(IP)(H 2 O)] to the selected enzyme.To validate the docking procedure, the co-crystalized ligand in the EGFR tyrosine kinase receptor (PDB Code: 1M17( was re-docked in its binding pocket.The co-crystalized ligand, 4-anilinoquinazoline (AQ4), revealed good poses with RMSD values below 2.0 Å and a high docking score (Table

Fig. 14 .
Fig. 14.Control cells of the three investigated cell lines.

Fig. 15 .
Fig. 15.Effect of Zn-MOF on the three investigated cell lines at different concentrations.

Zn(IP)(H 2 O)] at
a concentration of 10 µg/mL was used.The reaction solution was incubated at 37 ℃ for 15 min, followed by heating at 70 ℃ for 5 min, then the reaction mixture allowed to cool, and the developed turbidity was estimated spectrophotometrically at 660 nm.The control sample comprised a PBS and BSA in the absence of [

Zn(IP)(H 2 O)]. The
percentage of inhibition of denatured protein was calculated using the formula:where A 1 and A 2 represent the absorbance values of the control sample and the tested [

Table 2 .
The sharp peaks of the theoretical and experimental values of the prepared Zn-MOF.

Table 3 .
Physisorption properties of the Zn-MOF and its hybrid Ag/Zn-MOF.

Table 4 .
Ground state properties and atomic charges in terms of MPA and NPA of IPA linker and its Zn-MOF using B3LYP/6-311G and B3LYP/LANL2DZ respectively.

Table 8 .
Antimicrobial activities data in (mm) of Zn-MOF and its hybrid forms.NA No activity.Significant values are in bold.

Table 9 .
Previously reported Zn-MOFs' antimicrobial activity summary.was evaluated as a disrupting agent against the bacterial DNA using P. aeruginosa DNA as a representative example.The profile was verified by electrophoresis technology using the pure genomic DNA of P. aeruginosa pretreated with different concentrations of the tested compound.After a five-hour treatment, the tested [Zn(IP)(H 2 O)] at approximately 10 µg/mL caused fragmentation of P. aeruginosa genomic DNA, as observed on the gel (Fig.